In vitro evaluation of multiple antibacterial agents for the treatment of chronic staphylococcal anterior blepharitis.

OBJECTIVE: To evaluate the bactericidal efficacy of several compounds used in the treatment of chronic staphylococcal anterior blepharitis through an in vitro study. MATERIALS AND METHODS: Standard commercial strains of Staphylococcus aureus (SAu) (ATCC 25923 Culti-Loops) and coagulase-negative Staphylococcus (CoNS) (ATCC 12228 Culti-Loops) were cultured. Susceptibility tests were performed to vancomycin 30 µg, netilmicin 30 µg, hypochlorous acid (HOCl) 0.01% (Ocudox™, Brill®), Melaleuca alternifolia leaf oil (MeAl) (Navyblef® Daily Care, NOVAX®) and 1% chlorhexidine digluconate (DGCH) (Cristalmina™, Salvat®) using the agar disk diffusion method (Rosco Neo-Sensitabs®). After 24 h, the induced halos were measured with automatic calipers. The results were analyzed using the EUCAST- and CLSI potency Neo-Sensitabs® guidelines. RESULTS: Vancomycin induced a halo of 22.37 mm and 21.81 mm in SAu and CoNS, respectively. Netilmicin produced halos of 24.45 mm in SAu and 32.49 mm in CoNS. MeAl induced halos of 12.65 mm in SAu and 15.83 mm in CoNS. A 12.11 mm halo was found in SAu and an 18.38 mm halo in CoNS using HOCl. DGCH produced halos of 26.55 mm and 23.12 mm in SAu and CoNS, respectively. CONCLUSION: Netilmicin and vancomycin demonstrated antibiotic activity against both pathogens, so they can be alternative rescue therapies to treat chronic staphylococcal blepharitis. DGCH has efficacy against both comparable to antibiotics, while HOCl and MeAl show less efficacy.

Effects of explant size on epithelial outgrowth, thickness, stratification, ultrastructure and phenotype of cultured limbal epithelial cells.

PURPOSE: Transplantation of limbal stem cells is a promising therapy for limbal stem cell deficiency. Limbal cells can be harvested from either a healthy part of the patient's eye or the eye of a donor. Small explants are less likely to inflict injury to the donor site. We investigated the effects of limbal explant size on multiple characteristics known to be important for transplant function. METHODS: Human limbal epithelial cells were expanded from large versus small explants (3 versus 1 mm of the corneal circumference) for 3 weeks and characterized by light microscopy, immunohistochemistry, and transmission electron microscopy. Epithelial thickness, stratification, outgrowth, ultrastructure and phenotype were assessed. RESULTS: Epithelial thickness and stratification were similar between the groups. Outgrowth size correlated positively with explant size (r = 0.37; P = 0.01), whereas fold growth correlated negatively with explant size (r = -0.55; P < 0.0001). Percentage of cells expressing the limbal epithelial cell marker K19 was higher in cells derived from large explants (99.1±1.2%) compared to cells derived from small explants (93.2±13.6%, P = 0.024). The percentage of cells expressing ABCG2, integrin ß1, p63, and p63α that are markers suggestive of an immature phenotype; Keratin 3, Connexin 43, and E-Cadherin that are markers of differentiation; and Ki67 and PCNA that indicate cell proliferation were equal in both groups. Desmosome and hemidesmosome densities were equal between the groups. CONCLUSION: For donor- and culture conditions used in the present study, large explants are preferable to small in terms of outgrowth area. As regards limbal epithelial cell thickness, stratification, mechanical strength, and the attainment of a predominantly immature phenotype, both large and small explants are sufficient.

Effect of Transportation on Cultured Limbal Epithelial Sheets for Worldwide Treatment of Limbal Stem Cell Deficiency.

Limbal stem cell deficiency can be treated with transplantation of cultured human limbal epithelial cells (LEC). It can be advantageous to produce LEC in centralized labs and thereafter ship them to eye clinics. The present study used transport simulations of LEC to determine if vigorous shaking during transport altered the viability, morphology and phenotype during a 4 day-long storage of LEC with a previously described serum-free storage method. Inserts with LEC cultured on amniotic membranes were sutured to caps inside air-tight containers with generous amounts of 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES)-buffered minimal essential medium (MEM). The containers were distributed among the following testing conditions: 6 hours with full containers, 36 hours with full containers, 36 hours with container three quarters full of medium, and 36 hours with container full of medium containing a shear-protecting agent (Pluronic-F68). Compared to stored, but non-transported controls, no statistically significant changes in viability and immunohistochemical staining were observed. The epithelial sheets remained intact. However, an air-liquid interface in the containers reduced the number of desmosomes and hemi-desmosomes compared to the controls. In conclusion, cultured LEC sheets appear to endure vigorous shaking for at least 36 hours if the container is full.

Neurotrophic keratitis after transscleral diode laser cyclophotocoagulation. / Queratitis neurotrófica posciclofotocoagulación transescleral con láser diodo.

OBJECTIVE: To study the relationship between treatment with diode laser transscleral cyclophotocoagulation and development a neurotrophic keratitis due to the damage of the sensitive corneal innervation. METHODS: A study was conducted on 5 eyes of 5 patients who were treated with diode laser transscleral cyclophotocoagulation and soon developed neurotrophic ulcers. Personal characteristics of the patients were collected, as well as refraction and risk factors for corneal hypoesthesia, and the parameters of the laser used in the surgery. RESULTS: It was found that the 5 patients had predisposing factors of corneal hypoesthesia prior to surgery (chronic use of topical beta blockers, surgery with corneal incisions, diabetes mellitus, or corneal dystrophies); however none had developed neurotrophic keratitis until the cyclophotocoagulation was performed. It also showed that 4 of them were highly myopic, and they all were treated with high laser parameters (with an average of 2880 mW for 3s at an average surface of 275°), triggering neurotrophic ulcers between 10 and 35 days after surgery. CONCLUSION: Neurotrophic keratitis is a rare complication that can occur after diode laser transscleral cyclophotocoagulation, secondary to the damage of the long ciliary nerves. The emergence of this disorder can be triggered by the existence of previous risk factors, including high myopia, thus it is important to respect the recommended treatment parameters to prevent the development of this disorder.

[De novo growth of a capillary hemangioma of the conjunctiva]. / Crecimiento de novo de un hemangioma capilar conjuntival.

CLINICAL CASE: A 22-year-old woman patient, diagnosed with an inclusion cyst of the conjunctiva in the nasal sector of the left eye, who after 2 shot/needle injections in the lesion came to our clinic with a dense subconjunctival hemorrhage in four quadrants and with severe pain. After excision biopsy, a capillary hemangioma of the conjunctiva was diagnosed. DISCUSSION: Conjunctival capillary hemangioma is mainly a benign lesion, asymptomatic and mostly congenital in origin, its progression or de novo growth is rare in adulthood.

Post-surgical visual outcome and complications in Boston type 1 keratoprosthesis.

OBJECTIVE: To describe the visual outcome of patients who underwent Boston type 1 keratoprosthesis (KPro1) implantation, and describe serious sight-threatening post-operative complications. METHODS: We performed an analysis of the clinical records of all patients who underwent Boston keratoprosthesis implantation (BKI)in our institution from May 2006 to February 2011. RESULTS: A total of 41 eyes of 37 patients were included in the final analysis, of whom 22 (59.45%) were male and 15 were (40.54%) female. The mean age was 56.44 years (range 2-90). The most frequent diagnoses were bullous keratopathy, autoimmune diseases, such as Stevens-Johnson syndrome (SJS)/Lyell syndrome (LS), and aniridic keratopathy. The mean number of previous keratoplasties (PK) was 2.36 (range 0-8), the mean number of previous non-PK surgeries was 1.58 (range 0-9). The mean follow-up time was 22.17 months (range 3-46). The mean best corrected visual acuity (BCVA) logMAR before surgery was 2.05 (range 1.10-2.52), and the mean best corrected visual acuity achieved after surgery was 1.16 (range 0.08- 2.70). The most frequent complication was the formation of retroprosthetic membrane (RPM) which appeared in 22 (53.65%) eyes. Of these, 6 (27.27%) appeared after another surgery. Fourteen (63.63%) RPM required treatment, an average of 1.71 (range 1-4) laser YAG applications were performed, and surgical membranectomy was performed in 3 patients. Eleven (26.82%) eyes showed chorioretinal adhesion problems, 6 (14.63%) of which occurred after follow-up of BKI surgery. Infectious complications occurred in 7 (17.07%) cases; 2 (4.87%) patients had infectious keratitis and 5 (12.19%) endophthalmitis. CONCLUSIONS: Visual function improved in most patients. Those with prior multiple ocular surgeries and alterations of systemic immunity such as SJS, LS, and diabetes mellitus are at increased risk for serious sight-threatening complications, such as RPM, chorioretinal detachment and infection. Nevertheless, we consider KPro as an effective alternative in patients with multiple ocular pathology and imminent risk of rejection of a new KP.

[Limbal stem cell deficiency following multiple intravitreal injections]. / Insuficiencia limbar secundaria a múltiples inyecciones intravítreas.

CASE REPORT: A 68 year-old male with idiopathic choroidal polypoidal vasculopathy received seven intravitreal injections of bevacizumab in the supero-nasal quadrant. He later developed a localized limbal stem cell deficiency which required limbal transplantation from the same eye. Two months after surgery, a slight improvement of visual acuity was noted, along with an intact ocular surface, clear cornea and decreased inflammation. DISCUSSION: Repeated surgical trauma on the limbus and surrounding areas may damage the limbal stem cells, giving rise to a corneal epitheliopathy due to iatrogenic limbal deficiency. Repeated intravitreal injections may be considered as one of the possible causes.